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Immunological Identification of Multiple α‐Like Subunits of the γ‐Aminobutyric Acid A Receptor Complex Purified from Neonatal Rat Cortex
Author(s) -
Sato Thomas N.,
Neale Joseph H.
Publication year - 1989
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1989.tb07400.x
Subject(s) - protein subunit , microbiology and biotechnology , biology , receptor , biochemistry , gel electrophoresis , affinity chromatography , antibody , immunoprecipitation , peptide sequence , polyacrylamide gel electrophoresis , enzyme , gene , immunology
Antibodies were prepared against a synthetic peptide corresponding to amino acid sequences 174‐203 of the bovine γ‐aminobutyric acid A (GABA A ) receptor α1‐subunit. The antibodies recognized this synthetic α1 ‐peptide, but failed to react with the homologous peptide sequence, 170‐199, of the bovine β l ‐subunit. On Western blots, anti‐α1‐subunit antibody recognized a 50‐kilodalton (kDa) protein in affinity‐purified receptor preparations from adult rat cortex and cerebellum. In receptor purified from neonatal cortex, the antiα1‐antibody reacted with 50‐kDa, 53‐54‐kDa, and 59‐kDa proteins. After digestion with endoglycosidase F, these three protein bands retained differing electrophoretic mobilities. The 50‐kDa and 59‐kDa subunits of affinity‐purified neonatal receptor, which were photoaffinity‐labeled with [ 3 H]fluni‐trazepam, were immunoprecipitated to different extents by α‐subunit antibody. These data suggest the existence in GA‐BA A receptor from neonatal cortex of three proteins (50 kDa, 53 kDa, and 59 kDa) which have immunological homology to αl‐subunit of bovine GABA A receptor. The presence of an α‐ and a β‐like subunit with similar mobility on sodium dodecyl sulfate‐polyacrylamide gel electrophoresis may account for the relatively high concentration of protein in the 53‐54‐kDa band which has been observed in receptor purified from neonatal cortex. The presence of multiple α‐like sub‐units may be related to the presence of a relatively high concentration of type IIGABA receptor in this tissue.